In man, suppressor lymphocytes which modulate the response to transplantation antigens in vitro have been identified in the mixed lymphocyte culture reaction (MLR). Cyclosporin A (CsA), a novel immunosuppressive drug currently in use to prevent graft rejection and graft-versus-host disease preferentially inhibits cytolytic lymphocyte induction while permitting full activation of suppressor cells in the MLR. The suppressor cells appear to be antigen specific and maintain specific antigen tolerance in vitro. The activation and amplification of the suppressor cells in the presence of CsA, appears to be independent of IL-2 production and associated with a limited number of activated T-helper cells. Initial studies have identified a T-helper cell and a soluble factor capable of amplifying suppressor cell function without concomitant amplification of cytotoxic T cells. Characterization of these suppressor cell amplifying mechanisms is of critical importance for understanding the complex circuitry involved in the regulation of suppressor T cells. The cellular T-helper cell system identified by its relative resistance to CsA and by expression of Ia antigens will be characterized for its antigen specificity and will be assessed for its ability to secrete IL-2 or other soluble factors capable of amplifying suppressor cell function in primary MLR. Furthermore, utilizing recently developed cloning techniques, attempts will be undertaken to clone out the T-helper cell responsible for amplifying suppressor cell function. Subsequent studies using mixtures of cloned T-helper cell populations will attempt to detemine if there is regulatory influences (both positive and negative) on the T helper subset which amplifies suppressor cell activity. In addition, the soluble factor found in supernatants from CsA treated MLR cultures which support the amplification of suppressor cells in a T-helper cell depleted MLR and the apparent clonal amplification of antigen specific suppressor cells from day 10-12 MLR will be assessed for its antigen specificity, its ability to cooperate with MHC mismatched responder cells, and determine if this active component is a differentiation factor or growth factor specific for suppressor cells. If this activity is a growth factor, this component will be utilized in an attempt to clone out alloantigen specific suppressor T cells. Additionally, the biochemical attributes of this component will be defined. These studies should define some of the cellular and subcellular mechanisms required for suppressor cell amplification.